Ear infection - Pathogenesis, Lab diagnosis
Pathogenesis of ear infection
Otitis externa can be caused by local trauma, the presence of foreign bodies, or excessive moisture. Anatomic and physiological abnormalities of the auditory tube can predispose individuals to develop otitis media.
The auditory tube is responsible for ventilating the middle ear, equilibrating air pressure with the external ear, and protecting the middle ear from nasal secretions. If any of these functions are compromised, the middle ear may be filled with fluid and infection may occur.
The change in pressure may allow potentially pathogenic bacteria present in the nasopharynx into the middle ear which results in an ear infection.
Lab diagnosis
The laboratory diagnosis of ear infection begins with the collection of samples/specimens.
Specimen
ear swab, ear discharge
internal ear aspirate collected by an otolaryngologist
mastoid swab (Actual bone is preferred)
Transport
Transport of ear infection samples should be done anaerobically.
Microscopy
Methenamine silver stains are used to stain and observe pathogens under a microscope.
Culture
Diagnosis of otitis media (ear infection) is not generally done by culture but diagnosis of otitis externa is done by cultivation
Media used
Media used for diagnosis of ear infection is as follows:
BA, MA, and CA are the cultural media of choice
for specimens obtained by tympanocentesis, or from patients with chronic otitis media or mastoiditis, anaerobic cultures should be used
Inoculation, incubation
The agar plates are inoculated and incubated as:
MA = incubated aerobically at 37°C for 18-24 hours
BA = incubated at 35-37°C for 18-24 hours in 5% CO2
CA = incubated at 35-37°C for 18-24 hours in 5% CO2
Colony observation, presumptive identification
Observation of MA
Pink colonies suggest lactose fermenters (LF) pale yellow colonies suggest non-lactose fermenters (NLF).
Observation of BA
0.2 cm slightly β- hemolytic colony => maybe Staphylococcus aureus
pin-head sized, α- hemolytic colony => maybe Streptococcus pneumoniae
2mm, β-hemolytic colony => maybe Streptococcus pyogenes, Streptococcus agalactiae
large flat hemolytic colony with pigment => Haemophilus influenzae
0.2cm non-hemolytic opaque colony => Moraxella cattaharalis
Observation of CA
small greyish colonies => may be H. influenzae
Confirmatory identification of the isolate
If Streptococcus penumoniae is suspected
gram negative diplococci
Optochin sensitivity test (5 micrograms) – sensitive [5mm zone of inhibition (ZOI) seen]
Bile solubility test - test differentiates S. pneumoniae (soluble) from α-hemolytic Streptococci (insoluble)
Quelling reaction - enlarged capsule (positive) for S. pneumoniae
If Staphylococcus aureus is suspected
Gram-positive cocci in clusters
coagulase test = positive (differentiates S. aureus from CoNs)
growth in Mannitol Salt Agar (MSA) – golden yellow color:
In MSA, S. aureus gives yellow-colored colonies. Phenol red is present as an indicator. When S. aureus metabolizes mannitol, acid is produced as a by-product. A change in pH changes color to yellow.
If Streptococcus pyogenes is suspected
Gram-positive cocci in chains
Bacitracin sensitivity testing => ZOI 10 mm (susceptible)
Hippurate hydrolysis test: Negative
PYR test: positive
If Streptococcus agalactiae is suspected
Gram-positive cocci in chains
CAMP test: positive
PYR test: negative
If Pseudomonas aeruginosa is suspected
Gram-negative long rods
pigment production
Catalase positive, Oxidase positive
Characteristics on BA:
β-hemolytic
spreading, a flat colony with serrated edge
metallic sheen, bluish-green, red, or brown pigment
Biochemical test:
IMVIC (Indole, Methyl red, Voges-Proskauer, Citrate) test: -ve, -ve, -ve, +ve
Urease = Negative
TSI = Alk/Alk,
H2S negative
Nitrate reduction = positive
If Moraxella cattarrhalis is suspected
Gram-negative oval diplococci
Tri-butyryl esterase test: positive
DNase test: positive
No acid from carbohydrate
If Haemophilus influenza is suspected
Gram-negative diplococci
Satellitism test / X and V factor test: positive